Influence of microenvironment on engraftment of transplanted β-cells

Pancreatic islet transplantation into the liver provides a possibility to treat selected patients with brittle type 1 diabetes mellitus. However, massive early β-cell death increases the number of islets needed to restore glucose homeostasis. Moreover, late dysfunction and death contribute to the poor long-term results of islet transplantation on insulin independence. Studies in recent years have identified early and late challenges for transplanted pancreatic islets, including an instant blood-mediated inflammatory reaction when exposing human islets to the blood microenvironment in the portal vein and the low oxygenated milieu of islets transplanted into the liver. Poor revascularization of remaining intact islets combined with severe changes in the gene expression of islets transplanted into the liver contributes to late dysfunction. Strategies to overcome these hurdles have been developed, and some of these interventions are now even tested in clinical trials providing a hope to improve results in clinical islet transplantation. In parallel, experimental and clinical studies have, based on the identified problems with the liver site, evaluated the possibility of change of implantation organ in order to improve the results. Site-specific differences clearly exist in the engraftment of transplanted islets, and a more thorough characterization of alternative locations is needed. New strategies with modifications of islet microenvironment with cells and growth factors adhered to the islet surface or in a surrounding matrix could be designed to intervene with site-specific hurdles and provide possibilities to improve future results of islet transplantation

Acute hypoxia induces apoptosis of pancreatic β-cell by activation of the unfolded protein response and upregulation of CHOP

The success of pancreatic β-cells transplantation to treat type 1 diabetes has been hindered by massive β-cell dysfunction and loss of β-cells that follows the procedure. Hypoxia-mediated cell death has been considered one of the main difficulties that must be overcome for transplantation to be regarded as a reliable therapy. Here we have investigated the mechanisms underlying β-cell death in response to hypoxia (1% O2). Our studies show that mouse insulinoma cell line 6 (Min6) cells undergo apoptosis with caspase-3 activation occurring as early as 2 h following exposure to hypoxia. Hypoxia induces endoplasmic reticulum stress in Min6 cells leading to activation of the three branches of the unfolded protein response pathway. In response to hypoxia the pro-apoptotic transcription factor C/EBP homologous protein (CHOP) is upregulated. The important role of CHOP in the apoptotic process was highlighted by the rescue of Min6 cells from hypoxia-mediated apoptosis observed in CHOP-knockdown cells. Culturing isolated pancreatic mouse islets at normoxia showed intracellular hypoxia with accumulation of hypoxia-inducible factor-1α and upregulation of CHOP, the latter one occurring as early as 4 h after isolation. Finally, we observed that pancreatic islets of type 2 db/db diabetic mice were more hypoxic than their counterpart in normoglycemic animals. This finding indicates that hypoxia-mediated apoptosis may occur in type 2 diabetes

VEGFR2 pY949 signalling regulates adherens junction integrity and metastatic spread

The specific role of VEGFA-induced permeability and vascular leakage in physiology and pathology has remained unclear. Here we show that VEGFA-induced vascular leakage depends on signalling initiated via the VEGFR2 phosphosite Y949, regulating dynamic c-Src and VE-cadherin phosphorylation. Abolished Y949 signalling in the mouse mutant Vegfr2Y949F/Y949F leads to VEGFA-resistant endothelial adherens junctions and a block in molecular extravasation. Vessels in Vegfr2Y949F/Y949F mice remain sensitive to inflammatory cytokines, and vascular morphology, blood pressure and flow parameters are normal. Tumour-bearing Vegfr2Y949F/Y949F mice display reduced vascular leakage and oedema, improved response to chemotherapy and, importantly, reduced metastatic spread. The inflammatory infiltration in the tumour micro-environment is unaffected. Blocking VEGFAinduced disassembly of endothelial junctions, thereby suppressing tumour oedema and metastatic spread, may be preferable to full vascular suppression in the treatment of certain cancer forms

Protective Unfolded Protein Response in Human Pancreatic Beta Cells Transplanted into Mice

Background: There is great interest about the possible contribution of ER stress to the apoptosis of pancreatic beta cells in the diabetic state and with islet transplantation. Methods and Findings: Expression of genes involved in ER stress were examined in beta cell enriched tissue obtained with laser capture microdissection (LCM) from frozen sections of pancreases obtained from non-diabetic subjects at surgery and from human islets transplanted into ICR-SCID mice for 4 wk. Because mice have higher glucose levels than humans, the transplanted beta cells were exposed to mild hyperglycemia and the abnormal environment of the transplant site. RNA was extracted from the LCM specimens, amplified and then subjected to microarray analysis. The transplanted beta cells showed an unfolded protein response (UPR). There was activation of many genes of the IRE-1 pathway that provide protection against the deleterious effects of ER stress, increased expression of ER chaperones and ERAD (ER-associated protein degradation) proteins. The other two arms of ER stress, PERK and ATF-6, had many down regulated genes. Downregulation of EIF2A could protect by inhibiting protein synthesis. Two genes known to contribute to apoptosis, CHOP and JNK, were downregulated. Conclusions: Human beta cells in a transplant site had UPR changes in gene expression that protect against the proapoptotic effects of unfolded proteins

In vivo Hypoxia and a Fungal Alcohol Dehydrogenase Influence the Pathogenesis of Invasive Pulmonary Aspergillosis

Currently, our knowledge of how pathogenic fungi grow in mammalian host environments is limited. Using a chemotherapeutic murine model of invasive pulmonary aspergillosis (IPA) and 1H-NMR metabolomics, we detected ethanol in the lungs of mice infected with Aspergillus fumigatus. This result suggests that A. fumigatus is exposed to oxygen depleted microenvironments during infection. To test this hypothesis, we utilized a chemical hypoxia detection agent, pimonidazole hydrochloride, in three immunologically distinct murine models of IPA (chemotherapeutic, X-CGD, and corticosteroid). In all three IPA murine models, hypoxia was observed during the course of infection. We next tested the hypothesis that production of ethanol in vivo by the fungus is involved in hypoxia adaptation and fungal pathogenesis. Ethanol deficient A. fumigatus strains showed no growth defects in hypoxia and were able to cause wild type levels of mortality in all 3 murine models. However, lung immunohistopathology and flow cytometry analyses revealed an increase in the inflammatory response in mice infected with an alcohol dehydrogenase null mutant strain that corresponded with a reduction in fungal burden. Consequently, in this study we present the first in vivo observations that hypoxic microenvironments occur during a pulmonary invasive fungal infection and observe that a fungal alcohol dehydrogenase influences fungal pathogenesis in the lung. Thus, environmental conditions encountered by invading pathogenic fungi may result in substantial fungal metabolism changes that influence subsequent host immune responses

A local glucose-and oxygen concentration-based insulin secretion model for pancreatic islets

<p>Abstract</p> <p>Background</p> <p>Because insulin is the main regulator of glucose homeostasis, quantitative models describing the dynamics of glucose-induced insulin secretion are of obvious interest. Here, a computational model is introduced that focuses not on organism-level concentrations, but on the quantitative modeling of local, cellular-level glucose-insulin dynamics by incorporating the detailed spatial distribution of the concentrations of interest within isolated avascular pancreatic islets.</p> <p>Methods</p> <p>All nutrient consumption and hormone release rates were assumed to follow Hill-type sigmoid dependences on local concentrations. Insulin secretion rates depend on both the glucose concentration and its time-gradient, resulting in second-and first-phase responses, respectively. Since hypoxia may also be an important limiting factor in avascular islets, oxygen and cell viability considerations were also built in by incorporating and extending our previous islet cell oxygen consumption model. A finite element method (FEM) framework is used to combine reactive rates with mass transport by convection and diffusion as well as fluid-mechanics.</p> <p>Results</p> <p>The model was calibrated using experimental results from dynamic glucose-stimulated insulin release (GSIR) perifusion studies with isolated islets. Further optimization is still needed, but calculated insulin responses to stepwise increments in the incoming glucose concentration are in good agreement with existing experimental insulin release data characterizing glucose and oxygen dependence. The model makes possible the detailed description of the intraislet spatial distributions of insulin, glucose, and oxygen levels. In agreement with recent observations, modeling also suggests that smaller islets perform better when transplanted and/or encapsulated.</p> <p>Conclusions</p> <p>An insulin secretion model was implemented by coupling local consumption and release rates to calculations of the spatial distributions of all species of interest. The resulting glucose-insulin control system fits in the general framework of a sigmoid proportional-integral-derivative controller, a generalized PID controller, more suitable for biological systems, which are always nonlinear due to the maximum response being limited. Because of the general framework of the implementation, simulations can be carried out for arbitrary geometries including cultured, perifused, transplanted, and encapsulated islets.</p

Parkinson’s disease mouse models in translational research

Animal models with high predictive power are a prerequisite for translational research. The closer the similarity of a model to Parkinson’s disease (PD), the higher is the predictive value for clinical trials. An ideal PD model should present behavioral signs and pathology that resemble the human disease. The increasing understanding of PD stratification and etiology, however, complicates the choice of adequate animal models for preclinical studies. An ultimate mouse model, relevant to address all PD-related questions, is yet to be developed. However, many of the existing models are useful in answering specific questions. An appropriate model should be chosen after considering both the context of the research and the model properties. This review addresses the validity, strengths, and limitations of current PD mouse models for translational research

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The genetic architecture of the human cerebral cortex

The cerebral cortex underlies our complex cognitive capabilities, yet little is known about the specific genetic loci that influence human cortical structure. To identify genetic variants that affect cortical structure, we conducted a genome-wide association meta-analysis of brain magnetic resonance imaging data from 51,665 individuals. We analyzed the surface area and average thickness of the whole cortex and 34 regions with known functional specializations. We identified 199 significant loci and found significant enrichment for loci influencing total surface area within regulatory elements that are active during prenatal cortical development, supporting the radial unit hypothesis. Loci that affect regional surface area cluster near genes in Wnt signaling pathways, which influence progenitor expansion and areal identity. Variation in cortical structure is genetically correlated with cognitive function, Parkinson's disease, insomnia, depression, neuroticism, and attention deficit hyperactivity disorder